Abstract Human allogeneic islet cell transplantation as a therapy for cure for type 1 diabetes can be significantly improved if the deleterious effects of indefinite immunosuppression, particularly with their direct toxicity to [unreadable] cells, can be eliminated. Therefore, simple and effective donor-specific tolerance induction in such autoimmune diabetic hosts would be highly desirable. We have recently developed a novel tolerance regimen using i.v. infusion of donor splenocytes that are treated with a chemical cross-linker termed 1- ethyl-3-(3'-dimethylaminopropyl)-carbodiimide, or ECDI, and found that in the absence of any immunosuppression, this regimen is highly efficient in inducing durable donor- specific tolerance in allogeneic islet cell transplantation in the chemically-induced (streptozotocin) diabetes model. Similarly, soluble self-antigens such as insulin chemically fixed to syngeneic antigen presenting cells by ECDI have been independently shown to induce robust self-tolerance in the autoimmune diabetes NOD model. However, considerable obstacles exist in translating this methodology to a model harboring both alloimmunity and autoimmunity, i.e. in allogeneic islet transplantation for diabetic NOD mice, a model that is highly clinically relevant to patients with type 1 diabetes receiving deceased donor islet transplantation. This application proposes to take an integrated approach to identify crucial cellular components and signaling pathways required for tolerance by this protocol, as well as critical differences that exist in the autoimmune NOD hosts responsible for the ineffective tolerance induction. The application further proposes to study tolerance in the context of novel routes for islet allograft delivery that are potentially clinically applicable in humans. The ultimate goal of this study is to engineer novel tolerance regimen that is tailored to the specifics of islet allograft delivery, and is efficient and clinically feasible for human allogeneic islet cell transplantation. Successful completion of the proposed study will have significant impact on therapeutic options for patients with type 1 diabetes.